The proposed research will examine the functional and adaptive interactions among genes involved in the metabolic pathways of eukaryotic cells. An experimental adaptation approach will be used, in which strong selective pressure will be applied to a specific metabolic requirement in large steady-state cultures of yeast. Whole-genome changes in the transcriptional responses of evolved strains will be analyzed, and the genetic events underlying the adaptations will be assayed.The chemostat provides a very good model for this process. In the chemostat, cells are limited in growth by a particular scarce nutrient. The cell that overcomes that limitation out-competes the remainder of the culture and rapidly sweeps through the population. By following the progress of these sweeps, we can better understand the mutational mechanisms that underlie genetic responses to strong selection. The proposed research will follow up on these results and examine the extent to which they generalize. Planned limitations include nitrogen, phosphate and amino acids. The results of genome-wide gene-expression will be analyzed with yeast DNA microarrays. Gross chromosomal changes will be assayed using array comparative genomic hybridization (aCGH), and single point mutations found using genomic mismatch scanning (GMS). Data analysis will include standard statistical methods, including hierarchical clustering and singular value decomposition of expression data.